Muscular dystrophies show a progressive loss of muscle fibers and ultimate failure to regenerate muscle tissue. Cell-based therapy may restore muscle tissue, and adipose tissue may provide a facile source of cells for such therapy. We have been studying the mechanisms through which BMP or TGF-beta signaling can redirect pre-adipocyte differentiation and found that pre-adipocytes and primary adipose stromal cells can form muscle-like tissue when transplanted into immunodeficient mice. We propose to test the hypothesis that preadipocytes and adipose stromal cells can alter their differentiation to generate functional muscle in vivo, capable of contributing to tissue repair, and that BMP signaling regulates this differentiation. Smads are downstream effectors of signaling by TGF- beta and BMP and serve as cell-intrinsic regulators of mesenchymal differentiation, suggesting that manipulations of the activation levels of individual Smads will profoundly affect the differentiation of preadipocytes into muscle cells. In Aim 1 we will characterize the muscle-like cells generated from 3T3-F442A cells or human adipose stromal cells, and evaluate whether they arise from direct conversion or cell fusion with host myofibers. In Aim 2 we will determine whether transplanted adipogenic cells or adipocytes contribute to muscle repair in dystrophin-deficient nude mice, and determine whether engraftment improves muscle repair. We will also use a genetic approach to evaluate if endogenous (pre)adipocytes can differentiate into myocytes in response to injury and contribute to muscle (re)generation. In Aim 3 and future studies we will examine the role of Smad1 and Smad5, effectors of BMP signaling, and Smad3, effector of TGF-beta and myostatin signaling, as cell-intrinsic regulators of myogenic differentiation. These experiments should show us how to manipulate the myogenic differentiation by modifying Smad signaling. Our studies will hopefully provide the basis for the use of autologous preadipocytes and adipose stromal cells, a readily available cell source, for muscle regeneration and repair. PUBLIC HEALTH RELEVANCE. Patients with muscular dystrophy or defects in muscle repair would benefit from a therapy whereby injected cells would convert into muscle tissue. We recently found that cells that are becoming fat cells can be redirected to become muscle cells. We propose to characterize and to enhance the potential of these cells to become muscle cells, and to explore how they can help in the repair of muscle injury and to counteract the defects associated with muscular dystrophy, using mouse models. This research could provide a basis for the use of one's own fat in therapy to improve muscle repair and regeneration.